The effect of travoprost on primary human limbal epithelial cells and the siRNA-based aniridia limbal epithelial cell model, in vitro.

Purpose: Haploinsufficiency of the PAX6 gene is the primary pathogenic mechanism underlying classical congenital aniridia. Notably, at least 50% of patients with this condition develop glaucoma. Prostaglandin analogues, such as travoprost, are widely used to lower intraocular pressure in this patient population.

The Plus End-Directed Microtubule (Kinesin-3 Family) Motor Protein KIF13B Is Associated with the Photoreceptor Synaptic Ribbon Complex.

Retinal ribbon synapses are continuously active chemical synapses. The eponymous synaptic ribbon is anchored to the active zone neurotransmitter release sites of ribbon synapses, recruits synaptic vesicles and guides ribbon-associated synaptic vesicles to the release sites. RIBEYE is the major protein component of synaptic ribbons.

Altered Bcl-2/Caspase signaling and hypoxia-induced apoptosis in primary human aniridia limbal stromal cells, in CoCl2 mediated hypoxic stress, in vitro.

Purpose: The aim of this study was to investigate apoptosis in primary aniridia limbal stromal cells (LSCs) and to assess changes in the expression of genes and proteins associated with the apoptotic pathway in response to cobalt chloride (CoCl2)-induced hypoxic stress, in vitro.

Methods: Primary human limbal stromal cells were isolated from the limbal region of both aniridia (AN-LSCs; n = 8) and healthy (LSCs; n = 8) donors. The cells were treated with 0 µM, 50 µM, and 75 µM CoCl2 for 48 hours.

Reciprocal effects of conditioned medium on gene and protein expression of limbal epithelial cells and limbal fibroblasts in congenital aniridia.

Congenital aniridia is marked by substantial inflammatory changes to the ocular surface. However the exact mechanisms of epithelial-stromal interaction are not fully understood. The purpose of this study was to investigate inflammatory cytokine expression in limbal epithelial cells and fibroblasts following exposure to each other's conditioned medium (CM).

Increased sensitivity of primary aniridia limbal stromal cells to travoprost, leading to elevated migration and MMP-9 protein levels, in vitro.

Purpose: In congenital aniridia, not only limbal epithelial cells but also limbal stromal cells may contribute to the development of aniridia associated keratopathy (AAK). Secondary glaucoma affects 50-75% of patients with congenital aniridia, and prostaglandin analogs are commonly used for conservative treatment. This study aimed to explore the effect of travoprost on corneal limbal stromal cells from healthy (LSCs) and congenital aniridia subjects (AN-LSCs), in vitro.

Effect of retinoic acid treatment on the retinoic acid signaling pathway in a human siRNA-based aniridia limbal epithelial cell model, in vitro.

Purpose: In the present study, we evaluate gene and protein expression levels in an in vitro siRNA-mediated PAX6 knockdown limbal epithelial cell (LEC) model after RA treatment. This study aims to investigate the direct effects of active RA products and their association with key regulators of the RA signaling pathway in siRNA PAX6 knockdown LECs, providing further insights into the potential role of RA signaling in AAK pathogenesis.

Methods: Primary human limbal epithelial cells (LECs) were subjected to siRNA-mediated PAX6 knockdown to mimic PAX6 deletion in congenital aniridia (n = 8).

Gene expression study in the siRNA based aniridia cell model and in primary aniridia limbal epithelial cells following duloxetine and ritanserin treatment.

Progressive aniridia associated keratopathy is worsening visual acuity of congenital aniridia subjects lifelong. Restoration of PAX6 expression in PAX6 haploinsufficient limbal epithelial cells could be one therapeutic option. In a previous study using aniridia-like CRISPR/Cas9 genome-edited corneal epithelial cells, the antipsychotic drugs duloxetine and ritanserin increased PAX6 mRNA and protein expression.

Effect of isolated keratin 3 knockdown on gene expression of primary limbal epithelial cells without and with inflammatory stimuli.

Purpose: Studies have shown that keratin 3 (KRT3) expression is reduced in paired box 6 (PAX6) haploinsufficient primary limbal epithelial cells (LECs). The downregulation of KRT3 expression due to PAX6 haploinsufficiency is likely a critical factor in the development and progression of aniridia associated keratopathy (AAK). In addition, the ocular surface of congenital aniridia patients exhibits an inflammatory environment.

Effect of MiRNA 204-5P Mimics and Lipopolysaccharide-Induced Inflammation on Transcription Factor Levels, Cell Maintenance, and Retinoic Acid Signaling in Primary Limbal Epithelial Cells.

MicroRNA-204-5p (miR-204-5p) is a critical regulator of differentiation, structural maintenance, and inflammation in limbal epithelial cells (LECs). This study examined the role of miR-204-5p in modulating the gene expression related to transcription factors, cell structure, extracellular matrix remodeling, and retinoic acid signaling under normal and lipopolysaccharide (LPS)-induced inflammatory conditions. Using qPCR, we analyzed the mRNA levels of FOSL2, FOXC1, Meis2, PPARγ, ABCG2, PTGES2, IL-1β, IL-6, KRT3, KRT12, MMP2, MMP9, RARA, RARB, RXRA, RXRB, CRABP2, RBP1, RDH10, ADH7, ADH1A1, FABP5, CYP1B1, and CYP26A1, while changes in protein levels were assessed via Western blot or ELISA.

Comprehensive Analysis of Congenital Aniridia and Differential Diagnoses: Genetic Insights and Clinical Manifestations.

Introduction: Congenital aniridia (CA) is a severe and complex disorder involving the entire eye, primarily characterized by iris anomalies alongside other clinical features that pose significant risks to vision. This study seeks to offer a comprehensive overview of CA by detailing its clinical presentations, genetic underpinnings, associated phenotypes, and differential diagnoses. Additionally, it proposes a diagnostic framework to distinguish CA from other conditions that present with similar iris abnormalities.

Early Synapse-Specific Alterations of Photoreceptor Mitochondria in the EAE Mouse Model of Multiple Sclerosis.

Multiple sclerosis (MS) is an inflammatory autoimmune disease of the central nervous system (CNS) linked to many neurological disabilities. The visual system is frequently impaired in MS. In previous studies, we observed early malfunctions of rod photoreceptor ribbon synapses in the EAE mouse model of MS that included alterations in synaptic vesicle cycling and disturbances of presynaptic Ca homeostasis.

The cellular response of lipopolysaccharide-induced inflammation in keratoconus human corneal fibroblasts to RB-PDT: Insights into cytokines, chemokines and related signaling pathways.

Purpose: Rose Bengal Photodynamic Therapy (RB-PDT) offers dual therapeutic benefits by enhancing corneal stiffness and providing antibacterial activity, presenting significant potential for patients with keratoconus complicated by keratitis. Our purpose was to assess the effect of rose bengal photodynamic therapy (RB-PDT) on the expression of pro-inflammatory cytokines and chemokines, as well as on extracellular matrix (ECM)-related molecules, in lipopolysaccharide (LPS)-induced inflammation of keratoconus human corneal fibroblasts (KC-HCFs). Additionally, the involvement of the mitogen-activated protein kinase (MAPK) and nuclear factor kappa B (NF-κB) signaling pathways which are downstream of the Toll-like receptor 4 (TLR4) pathway were examined.

Expression of PAX6 and Keratocyte-Characteristic Markers in Human Limbal Stromal Cells of Congenital Aniridia and Healthy Subjects, .

Purpose: Our aim was to examine the expression of PAX6 and keratocyte-specific markers in human limbal stromal cells (LSCs) in congenital aniridia (AN) and in healthy corneas, .

Methods: Primary human LSCs were extracted from individuals with aniridia (AN-LSCs) ( = 8) and from healthy corneas (LSCs) ( = 8). The cells were cultured in either normal-glucose serum-containing cell culture medium (NGSC-medium) or low-glucose serum-free cell culture medium (LGSF-medium).

miRNA Expression Profile in Primary Limbal Epithelial Cells of Aniridia Patients.

Purpose: This study evaluates the microRNA (miRNA) expression profile in primary limbal epithelial cells (pLECs) of patients with aniridia.

Methods: Primary human LECs were sampled and isolated from 10 patients with aniridia and 10 healthy donors. The miRNA profile was analyzed using miRNA microarrays.

Systemic Diseases in Patients with Congenital Aniridia: A Report from the Homburg Registry for Congenital Aniridia.

Introduction: Congenital aniridia is increasingly recognized as part of a complex syndrome with numerous ocular developmental anomalies and non-ocular systemic manifestations. This requires comprehensive care and treatment of affected patients. Our purpose was to analyze systemic diseases in patients with congenital aniridia within the Homburg Aniridia Registry.

Effect of Ritanserin and Duloxetine on the Gene Expression of Primary Aniridia and Healthy Human Limbal Stromal Cells, In Vitro.

Introduction: In congenital aniridia caused by mutations in paired box 6 (PAX6), PAX6 influences the migration and differentiation of limbal epithelial cells (LECs), thereby playing a pivotal role in aniridia-associated keratopathy. The antidepressants ritanserin and duloxetine affect PAX6 expression in LECs. Limbal stromal cells, which support limbal epithelial stem cells, are crucial in the limbal stem cell niche.

Rose Bengal Photodynamic Therapy (RB-PDT) Modulates the Inflammatory Response in LPS-Stimulated Human Corneal Fibroblasts By Influencing NF-κB and p38 MAPK Signaling Pathways.

Purpose: To investigate the effect of rose bengal photodynamic therapy on lipopolysaccharide-induced inflammation in human corneal fibroblasts. Furthermore, to analyze potential involvement of the mitogen-activated protein kinase and nuclear factor kappa B signaling pathways in this process.

Methods: Human corneal fibroblast cultures underwent 0-2.

A Novel Cre Recombinase Mouse Strain for Cell-Specific Deletion of Floxed Genes in Ribbon Synapse-Forming Retinal Neurons.

We generated a novel Cre mouse strain for cell-specific deletion of floxed genes in ribbon synapse-forming retinal neurons. Previous studies have shown that the RIBEYE promotor targets the expression of recombinant proteins such as fluorescently tagged RIBEYE to photoreceptors and retinal bipolar cells and generates fluorescent synaptic ribbons in situ in these neurons. Here, we used the same promotor to generate a novel transgenic mouse strain in which the RIBEYE promotor controls the expression of a Cre-ER(T2) recombinase (RIBEYE-Cre).

Ciliary Proteins Repurposed by the Synaptic Ribbon: Trafficking Myristoylated Proteins at Rod Photoreceptor Synapses.

The Unc119 protein mediates transport of myristoylated proteins to the photoreceptor outer segment, a specialized primary cilium. This transport activity is regulated by the GTPase Arl3 as well as by Arl13b and Rp2 that control Arl3 activation/inactivation. Interestingly, Unc119 is also enriched in photoreceptor synapses and can bind to RIBEYE, the main component of synaptic ribbons.

An easy, fast and "low-tech"-equipment-requiring alternative method to optimize immunolabelling conditions for pre-embedding immunogold electron microscopy and to correlate light and electron microscopical immunogold labelling results.

Correlating light microscopic immunolabelling results with electron microscopic data is of great interest in many fields of biomedical research but typically requires very specialized, expensive equipment and complex procedures which are not available in most labs. In this technical study, we describe an easy and "low-tech"-equipment-requiring pre-embedding immunolabelling approach that allows correlation of light microscopical immunolabelling results with electron microscopic (EM) data as demonstrated by the example of immunolabelled synaptic ribbons from retinal rod photoreceptor synapses. This pre-embedding approach does not require specialized embedding devices but only commonly available equipment.